Small RNA sequencing analysis identified miR159a as a novel candidate for activity in plant-derived nanovesicles from limon, hassaku, and sudachi.

Owing to their safety, plant-derived nanovesicles (PDNVs), particularly those derived from edible plants, are expected to be natural delivery carriers for microRNAs (miRNAs). Among PDNVs, Citrus limon-derived extracellular vesicle-like nanovesicles have been shown to exert antitumor effects. We previously reported that PDNVs derived from C. limon L. exert selective inhibitory effects on the proliferation of p53-inactivated colon cancer cells. However, miRNAs in C. limon L.-derived nanovesicles have not yet been identified, and it remains unclear which miRNAs are contained in these PDNVs. We herein extracted PDNVs from C. limon L., C. hassaku, and C. sudachi and identified and compared the miRNAs present within them via next-generation sequencing. A total of 158 miRNAs were identified, of which 109 were known miRNAs and 49 were novel miRNAs. Comparisons of miRNAs expressed by C. limon L., C. hassaku, and C. sudachi revealed 77 miRNAs that were commonly expressed. Among all the libraries, miR159a had the highest number of reads, and the same result was obtained when it was converted to TPM. The present results raise the possibility that miR159a could be involved in the biological properties of citrus‑derived nanovesicles, given that it was the most abundantly expressed miRNA.